Synthesis and biological properties of oxazolinodaunorubicin--a new derivative of daunorubicin with a modified daunosamine moiety.

Oxazolinodaunorubicin, a new daunorubicin derivative with a modified daunosamine moiety, was synthesized. The biological properties of this derivative and the parent daunorubicin were compared. The results showed antiproliferative activity of the derivative with significantly lower toxicity (an LD(50) value ca. 20 times higher than that of parent daunorubicin) and an ability to completely overcome the resistance of cancer cells to this drug in vitro. Cardiotoxicity determination using male mice treated with a single dose of 75% of the LD(50) value indicated that the cardiotoxicity of new analog was much lower than that of the parent drug. Preliminary results in transplanted murine tumor models revealed that a single-dose injection of the tested compounds exhibited antitumor activity in P388 and L1210 leukemia and 16/C mammary adenocarcinoma bearing mice.

Antitumor effect of combined treatment of mice with cytostatic agents and bacteriophage T4.

The past few years have shown significant resurgent interest in the old concept of bacteriophage therapy. Some research groups continue to develop whole bacteriophage preparations as alternatives to antibiotic antibacterial treatment. However, improvements in the methods of purification of phage preparations open new opportunities in the successful treatment of antibiotic-resistant bacterial infections. An open question remains on whether bacteriophage preparations (BP) can be safely applied in antibacterial treatment of patients suffering from infections as a consequence of immunosuppression caused by anticancer chemotherapy. The aim of this study was to evaluate the potential modulating effect of bacteriophage T4 preparations administered to mice bearing s.c. or i.v. inoculated B16 melanoma and treated with conventional anticancer drugs, i.e. cyclophosphamide (CY), cisplatin (CPt) or 5-fluorouracil (5-FU). Treatment of mice with (BPT) T4 preparation slightly potentiated the antimetastatic effect of CY. Importantly, no combination of phage-cytostatic treatment resulted in a decrease in the antimetastatic or antitumour effects of an applied drug. This suggests the possibility of safe combination of bacteriophage preparations with popular antitumour drugs.

Liposomal formulation of 5-fluorocytosine in suicide gene therapy with cytosine deaminase--for colorectal cancer.

It is generally accepted that successful gene therapy depends on two major factors: tumor-specific expression of a therapeutic gene and the efficient transfer of a therapeutic gene to tumor cells. For gene-directed enzyme prodrug therapy (GDEPT) involving Escherichia coli cytosine deaminase (CD) and 5-fluorocytosine (5-FC), several tumor-specific promoters and virus-based vectors were used. No attention whatsoever was paid to the way of 5-FC delivery to solid tumors, despite the fact that the delivery of drugs to such tumors is generally low because of their insufficient transfer from the blood. To compare the effectiveness of GDEPT with free and liposomal 5-FC, the prodrug was encapsulated in liposomes composed of dipalmitoylphosphatidylcholine (DPPC) and cholesterol (1:1). When the liposomal form of 5-FC was administered i.v., mice treated with a dose of 5mg of liposomal 5-FC/kg body weight for 10 days, showed complete regression of transplanted tumors and complete cure was observed, whereas in animals treated with the same amounts of the free prodrug, 50% tumor regression and only insignificantly prolonged median survival were found. In summary, these results showed a remarkable enhancement of the antitumor effects of the liposomal form of 5-FC in comparison with the free prodrug. Therapy with liposomal 5-FC thus represents a new approach to achieving a high local concentration of the prodrug for suicide gene therapy using E. coli CD.

Antitumor activity of mannan-methotrexate conjugate in vitro and in vivo.

Conjugation of anticancer drugs with different carriers has been extensively studied recently as a potential method of obtaining improved drug forms. The conjugation often results in the increase of the therapeutic effect, alteration of a toxicity profile, and/or selective targeting of therapeutic agent to the tissue of interest. We have synthesized mannan-methotrexate conjugate by means of methotrexate anhydride and studied its antitumor properties both in vitro and in vivo in comparison with free methotrexate. Mannan-methotrexate conjugate showed significantly improved antitumor activity compared to free methotrexate in the model of P388 mouse leukemia disseminated in the peritoneal cavity treated with intraperitoneally injected chemotherapy. Conversely, the antitumor effects of free methotrexate and mannan-methotrexate conjugate were comparable when leukemia was implanted subcutaneously and chemotherapy agents were administered intravenously. These results suggest that mannan-methotrexate conjugate should be further investigated as a potential therapeutic agent for intraperitoneally disseminated tumors.

Synthesis and in vitro antiproliferative activity of new 1,3-(oxytetraethylenoxy)-cyclotriphosphazene derivatives.

A new series of 1,3-(oxytetraethylenoxy)-cyclotriphosphazene derivatives containing aziridine or salicylaldehyde (2-hydroxybenzaldehyde) or its Schiff base units after condensation with 2-chloroethylamine and anthraquinone groups as co-substituents has been synthesized. Their cytostatic activity in vitro against the HL-60, A549 and HCV29T cancer cell lines has been studied. Some of the compounds exhibited antiproliferative activity in the range of the international criteria for synthetic agents (4 microg/ml) against the cell lines being tested.

Biological properties of new derivatives of daunorubicin.

In the search for new derivatives of daunorubicin with high activity and/or the ability to overcome the drug resistance barrier of cancer cells, some new analogs of amidino-daunorubicin, containing the chiral substituent in the formamidine group (-N=CH-N<) at the C-3' position of daunosamine moiety, have been synthesized. In order to estimate the influence of the configuration of the chiral group on the biological properties of the new derivatives of daunorubicin, three chiral amines, namely 1-cyclohexyl-ethylamine, 1-phenylethylamine and N-methyl-l-phenyl-ethylamine, both R and S isomers and their racemates, were used. These new compounds were tested for their cytotoxic activity in vitro against the cells of A549, SW707, T47D and HCV29T cancer lines. The resistance index (RI) values were obtained using the cells of the sensitive LoVo, MES-SA, HL-60 human cancer cell lines, as well as their resistant sublines (LoVo/Dx, MES-SAIDX5 and HL-60/MX2, respectively). All obtained derivatives appeared to be able to overcome the drug resistance barrier of cancer cells.

Antitumor properties of diastereomeric and geometric analogs of vitamin D3.

Analogs of 1,25-dihydroxyvitamin D3 with a reversed configuration at C-1 or C-24 and E or Z geometry of the double bond at C-22 in the side chain or at C-5 in the triene system were examined for their antiproliferative activity in vitro against a spectrum of various human cancer cell lines. The analogs coded PRI-2201 (calcipotriol), PRI-2202 and PRI-2205, such as calcitriol and tacalcitol (used as a referential agents), revealed antiproliferative activity against human HL-60, HL-60/MX2, MCF-7, T47D, SCC-25 and mouse WEHI-3 cancer cell lines. The toxicity studies in vivo showed that PRI-2202 and PRI-2205 are less toxic than referential agents. Even at total doses of 2.5-5.0 mg/kg distributed during 5 successive days, no changes in body weight were observed. Calcitriol and tacalcitol showed toxicity in the same protocol at 100 times lower doses. Calcipotriol was lethal to all mice after administration of a total dose of 5.0 mg/kg. The analog PRI-2205 appeared to be more active in mouse Levis lung cancer tumor growth inhibition than calcitriol, calcipotriol or PRI-2202. This analog did not reveal calcemic activity at doses which inhibit tumor growth in vivo nor at higher doses.

Hoc protein regulates the biological effects of T4 phage in mammals.

We previously investigated the biological, non-antibacterial effects of bacteriophage T4 in mammals (binding to cancer cells in vitro and attenuating tumour growth and metastases in vivo); we selected the phage mutant HAP1 that was significantly more effective than T4. In this study we describe a non-sense mutation in the hoc gene that differentiates bacteriophage HAP1 and its parental strain T4. We found no substantial effects of the mutation on the mutant morphology, and its effects on electrophoretic mobility and hydrodynamic size were moderate. Only the high ionic strength of the environment resulted in a size difference of about 10 nm between T4 and HAP1. We compared the antimetastatic activity of the T2 phage, which does not express protein Hoc, with those of T4 and HAP1 (B16 melanoma lung colonies). We found that HAP1 and T2 decreased metastases with equal effect, more strongly than did T4. We also investigated concentrations of T4 and HAP1 in the murine blood, tumour (B16), spleen, liver, or muscle. We found that HAP1 was rapidly cleared from the organism, most probably by the liver. Although HAP1 was previously defined to bind cancer cells more effectively (than T4), its rapid elimination precluded its higher concentration in tumours.

Antiproliferative activity of vitamin D compounds in combination with cytostatics.

Calcitriol is a potent antiproliferative agent against various tumour cells in vitro. Here, the results of a study on vitamin D compounds (calcitriol's analogues PRI-1906 and PRI-2191) as potential agents in combined antitumour therapy in vitro are presented. Applying antiproliferative SRB and MTT assays, the growth inhibitory effects of the vitamin D compounds, applied alone or in combination with either cisplatin or doxorubicin, were measured. The following cancer cell lines were employed: A549 (human non-small cell lung carcinoma), B16 (murine melanoma), CCRF, HL-60 (human leukaemia), SW707 (human colon cancer), MCF-7, T47D (human breast cancer), WEHI-3 (mouse leukaemia) and normal cells: BALB 3T3 (normal murine fibroblast cell line). It was shown that the treatment of tumour cells, which are sensitive to vitamin D compounds, with the combination of vitamin D compounds and cytostatics decreased the inhibitory concentration 50% (IC50) values compared with the effects of the cytostatics applied alone.

Key factors in experimental mouse hematopoietic stem cell transplantation.

The first mouse model of hematopoietic stem cell transplantation (HSCT) was developed more than 50 years ago. HSCT is currently being widely used in a broad range of research areas, which include studies of the engraftment process, the pathogenesis of graft-versus-host disease and possible ways of its treatment and prophylaxis, attempts to use the graft-versus-leukemia/tumor effect in treating hematological and oncological malignancies, cancer vaccine development, induction of transplanted organ tolerance, and gene therapy. However, although this model is widely distributed, many laboratories use different protocols for the procedure. There are a number of papers discussing different HSCT protocols in clinical work, but no articles summarizing mouse laboratory models are available. This review attempts to bring together different details about HSCT in the mouse model, such as the types of transplantation, possible pretreatment regimens and their combinations, methods and sources of graft harvesting and preparation for the transplantation procedure, the influence of graft cell dose and content on the engraftment process, the transplantation method itself, possible complications, symptoms and techniques of their prophylaxis or treatment, as well as follow-up and engraftment assessment. We have also tried to reflect current knowledge of the biology of the engraftment.

Effect of structural modifications of anthracyclines on the ability to overcome drug resistance of cancer cells.

In the search for new derivatives of anthracycline antibiotics with the ability to overcome the drug resistance barrier, a series of new analogs of these antibiotics, containing the amidino group at C-3' position of the daunosamine moiety, have been synthesized. The new compounds were tested for their cytotoxic activity in vitro against the sensitive LoVo, MES-SA and HL-60 human cancer cell lines as well as their resistant sublines: LoVo/Dx, MES-SA/Dx5 and HL-60/MX2, respectively. The majority of these derivatives appeared to be able, completely or partially, to overcome the drug resistance barrier of cancer cells. The effect of structural modification on this ability was determined. The obtained results indicated that introduction of the amidino group into the daunosamine moiety of anthracycline molecules appears to overcome the drug resistance of cancer cells.

The effect of the substitution level of some dextran-methotrexate conjugates on their antitumor activity in experimental cancer models.

Methotrexate (MTX) is widely used in the treatment of a number of oncological and hematological diseases. Due to its known limitations, MTX is often conjugated with different carriers to obtain amended forms of the drug. In this study, the potential influence of the substitution level (loading ratio) of the dextran T10- and T40-based MTX conjugates (D-MTX) on their properties were investigated in vitro and in vivo. The clear dependence of the in vitro antiproliferative effect on the substitution level was established only in the case of the dextran T10-based preparations (T10-MTX conjugates). Conjugates with the higher substitution level had the lower antiproliferative effect. For the dextran T40-based (T40-MTX conjugates) set no similar relationship was observed in the tested range of substitution levels, nor was any dependence observed between the biological properties of the D-MTX preparations in vivo and their substitution levels. However, the difference between the two conjugates was well pronounced in a multiple-dose schedule, when the advantage of T40-MTX over T10-MTX was cumulative during the prolonged course of administration.

Antitumor properties and toxicity of dextran-methotrexate conjugates are dependent on the molecular weight of the carrier.

Methotrexate (MTX) is widely utilized in the clinical treatment of many forms of cancer. However, the drug has a short plasma half-life and causes toxic effects on normal proliferating cells. Conjugation with carriers is a possible way to alter these disadvantageous pharmacokinetics. Our aim was to synthesize dextran-MTX (D-MTX) conjugates, using carriers with molecular weights (Mw) ranging from 10 kDa to 500 kDa. Their in vitro and in vivo properties were compared with free MTX. The in vitro studies revealed that D-MTX conjugates had 4- to 10-fold lower antiproliferative effects against neoplastic cell lines compared to free MTX. There was a negative relationship between the Mw of the carrier and the antiproliferative effect of the respective conjugate. The data obtained in a mouse leukemia P388 in vivo model suggested that a lower in vitro antiproliferative effect of the conjugates does not result in diminished antileukemic activity in vivo. The toxicity of the conjugates was greater in comparison with the parent drug and tended to rise with increasing Mw. However, no superiority over free MTX in terms of an antileukemic effect was demonstrated. In particular, the D-MTX conjugate based on the dextran with Mw 10 kDa showed a comparable antileukemic effect with an even lower toxicity than that of free MTX. The data suggest that at least the toxicity of conjugates is dependent on the Mw of the carrier. This fact should be taken into account when designing new anticancer polymer-drug compounds.

Synthesis and antiproliferative activity of some 5-substituted 2-(2,4-dihydroxyphenyl)-1,3,4-thiadiazoles.

A series of new 5-substituted 2-(2,4-dihydroxyphenyl)-1,3,4-thiadiazoles has been synthesised and evaluated for their antiproliferative activity. The compounds were prepared by the reaction of the sulphinylbis(2,4-dihydroxythiobenzoyl) (STB) wit hydrazides or carbazates. The panel substitution included alkyl, alkoxy, aryl and heteroaryl derivatives. The structures of compounds were identified from the elemental, IR, (1)H NMR and MS spectra analysis. The highest antiproliferative activity against the cells of human cancer lines for 2-(2,4-dihydroxyphenyl)-5-(4-methoxybenzyloxy)-1,3,4-thiadiazole was found with ID(50) values comparable (HCV29T and SW707) or significantly lower (T47D) than for cisplatin applied as the reference compound. The influence of 5-substiution type of 2-(2,4-dihydroxyphenyl)-1,3,4-thiadiazoles on antiproliferative activity is discussed.

Synthesis and antiproliferative activity of N-substituted 2-amino-5-(2,4-dihydroxyphenyl)-1,3,4-thiadiazoles.

A number of N-substituted 2-amino-5-(2,4-dihydroxyphenyl)-1,3,4-thiadiazoles were synthesized and evaluated for their antiproliferative activities. The panel substitution included alkyl, aryl, and morpholinoalkyl derivatives. The structures of compounds were identified from elemental, IR, (1)H NMR, (13)C NMR and MS spectra analyses. The cytotoxicity in vitro against the four human cell lines: SW707 (rectal), HCV29T (bladder), A549 (lung), and T47D (breast) was determined. Alkyl and morpholinoalkyl derivatives exhibited significantly lower effect than phenyl ones. The highest antiproliferative activity was found for 2-(2,4-dichlorophenylamino)-5-(2,4-dihydroxyphenyl)-1,3,4-thiadiazole, with ID(50) two times lower (SW707, T47D) than for cisplatin studied comparatively as the control compound.

Synthesis and in vitro antileukemic activity of some new 1,3-(oxytetraethylenoxy)cyclotriphosphazene derivatives.

A new series of 1,3-(oxytetraethylenoxy)cyclotriphosphazene derivatives bearing 2-chloroethylamine or salicylaldehyde (2-hydroxybenzaldehyde) or its Schiff base (after condensation with 2-chloroethylamine) units and having also 2-naphthyl or anthraquinone groups as cosubstituents has been synthesized. The in vitro cytotoxic activity of these compounds against a panel of four cancer cell lines has been studied. Most of the compounds exhibited antiproliferative activity in the range of the international criterion for synthetic agents (4 microg/mL) against the MOLT4, L 1210, HL-60, and P388 cell lines chosen for testing.

Synthesis and antiproliferative activity in vitro of new 2-, 3- or 4-substituted pyrido[2',3':3,4]pyrazolo[1, 5-a]pyrimidines.

The synthesis of several new 2-, 3- or 4-substituted pyrido[2',3':3,4]pyrazolo[1,5-a]pyrimidines is described. The obtained compounds were tested for their antiproliferative activity in vitro. Two of them: 3-chloro-2-methylpyrido[2',3':3,4]pyrazolo[1,5-a] pyrimidin-4-one [X] and 2,3-cyclopentylpyrido [2',3':3,4] pyrazole[1,5-a]pyrimidin-4-one [XII] revealed weak cytotoxic activity against the cells of human bladder cancer cell lines: LoVo, MCF-7, MES-SA and HCTV29T. The structures of the products II - XII were established on the basis of elemental analysis and spectral data (IR, 1H NMR and MS).

[TRPM7: a protein responsible for magnesium homeostasis in a cell]. / TRPM7--bialko odpowiedzialne za homeostaze magnezu w komórce.

Magnesium is an important cofactor in biological processes. For many years it has been considered that magnesium homeostasis in a cell is regulated by its eflux from a cell and not by its influx. It has also been considered that the Na+/Mg2+ antiport plays the main role. In recent years, many experiments have been carried out to understand the mechanisms of Mg2+ transport in an organism. These experiments have led to some new conclusions. It was confirmed that the level of magnesium in a cell is probably also regulated by its influx to the cell. One of the last scientific findings is the discovery of the TRPM (transient receptor potential melastatin) protein family. TRPM6 and TRPM7, bifunctional proteins with kinase and ion channel activities, are responsible for magnesium homeostasis. The discovery of these proteins led to a better understanding of magnesium homeostasis. It was confirmed that TRPM6 protein is responsible for homeostasis in the whole organism and that TRPM7 may regulate the level of magnesium in the cell. TRPM7 also has other functions. One of those newly recognized is the phosphorylation of annexin 1. However, many activities and functions of TRPM7 have not yet been described. This paper is a review of knowledge of TRPM7 transmembrane protein, which is responsible for the magnesium homeostasis in the cell. It briefly presents the main functions and structure of TRPM7. It also describes the mechanisms of its biological activity.

Synthesis of 5,6-dimethyl-9-methoxy-1-phenyl-6H-pyrido[4,3-b]carbazole derivatives and their cytotoxic activity.

Starting from 2-(6-methoxy-1-methyl-9H-carbazol-2-yl)ethylamine 7 and mixed anhydrides of 4-nitrobenzoic acid or 4-methoxybenzoic acid, the corresponding 5,6-dimethyl-9-methoxy-1-(4-substituted phenyl)-6H-pyrido[4,3-b]carbazoles 11a-b, 5,6-dimethyl-9-hydroxy-1-(4-substituted phenyl)-6H-pyrido[4,3-b]carbazoles 12a, 12c, and their quaternary salts 13a-d were obtained. The four new pyridocarbazole derivatives 12a-c and 13d satisfy the international activity criterion for synthetic compounds, namely an ID(50) value lower then 4 microg/mL in preliminary in vitro cytotoxic activity screening against the A549 cell line (non-small cell lung cancer).

Influence of the structure of new anthracycline antibiotics on their biological properties.

In the search for new derivatives of anthracycline antibiotics with advantageous biological properties, particularly with lower toxicity and/or higher activity, a series of new analogs of antibiotics applied in therapy such as daunorubicin, doxorubicin, as well as epidoxorubicin and, for comparison, analogs of epidaunorubicin, have been synthesized. Our results show that the new derivatives have antiproliferative activities similar to or higher than the parent antibiotics. The toxicities of these analogs were significantly lower, with LD50 values from 1.8- to 18.4-fold higher than the referential drugs. Cardiotoxicity determinations, using male mice treated with a single dose of 75% of the LD50 values of all tested compounds, indicated that the cardiotoxicity of the new analogs is significantly lower than that of the parent drugs.